The proposed investigation is designed to attack specific questions on the nature, specificity and biological function of DNA modification. The relationship of DNA methylation and chromatin structure will be studied in the protozoan, Tetrahymena. The sequence specificity of the Tetrahymena DNA-adenine methylase will be elucidated. We will assess the possibility that methylation is involved as a signal for nicking/gapping of macronuclear DNA within the tandem repeat (C-C-C-C-A-A)n. We will attempt to develop an in vitro assay system to screen purification of the phage Mu mom plus protein; this protein is responsible for an unusual (sequence-specific) modification of adenine residues in phage Mu DNA. We will determine whether host-specific dam plus DNA methylation is required for transcription of the Mu beta-region (location of the mom plus gene). We will examine the question of whether N to the sixth power-methyladenine (a normal methylated base in most prokaryote DNAs) is mutagenic. We will clone two alternative forms of the phage T4 dam plus and dam h genes into E. coli dam minus and determine the effects on mutation rate. We will also explore the possibility of cloning these genes into yeast and examining the genetic consequences of having an additional, foreign DNA modification pattern.